The long-term goals of this proposal are to understand the molecular and cellular basis for nicotine's addictive properties and to elucidate the functional role of neuronal nicotinic acetylcholine receptors (nAChRs) in the many centrally mediated behavioral and physiological effects of nicotine. This proposal focuses on alpha3-subunit-containing nAChRs which are found at high density in the medial habenula and which also have been implicated in the ventral tegmental area (VTA), a well recognized component of the reward pathway. The medial habenula is an evolutionarily conserved brain region that has been shown to be very sensitive to the neurotoxic effects of nicotine. Furthermore, it has been suggested that some of the cognitive deficits seen in schizophrenia may be linked to underlying pathology in the medial habenula. The main tool to be utilized in this study is a novel knock-in mouse in which the nicotinic alpha3-subunit has been replaced with one containing five amino acid substitutions that impart sensitivity to pharmacological blockade by alpha-bungarotoxin (Bgtx). This knock-in mouse enables the use of the classic pharmacological antagonist, Bgtx, to probe the functional and behavioral role of alpha3-containing nAChRs in the medial habenula and VTA. First, it will be important to confirm the regional expression of Bgtx-sensitive, alpha3- containing neuronal nAChRs in frozen sections of brains isolated from mice heterozygous for the targeted alpha3/alpha1[5] mutation (+/tm1.1). Autoradiography of radioactive-Bgtx binding sites will be used for this purpose. Following stereotaxic cranial cannulation, Bgtx will be microinjected into the medial habenula and VTA of (+/tm1.1) mice to determine the effect of pharmacological blockade of alphas-containing nAChRs on three nicotine-associated behaviors. These include: 1) the hypolocomotor effect of acute, low-dose systemic nicotine;2) entrained oral preference for nicotine;and 3) nicotine-induced seizures. Relevance: Nicotine is an extremely addictive drug responsible for up to 20% of all preventable mortality in the western world. Nicotine also significantly enhances cognitive performance, and some inherited forms of epilepsy involve nicotinic receptors. In addition, a loss of cholinergic neurons is implicated in Alzheimer's disease, a disorder currently lacking effective treatment. Understanding the molecular interactions of nicotine with its receptors in the central nervous system therefore has significant potential to benefit human health.